ABSTRACT/PROJECT SUMMARY: There is a strong relationship between sleep and Alzheimer?s disease (AD). Individuals with sleep apnea or disrupted sleep are at greater risk of developing AD. This is likely directly related, at least in part, to brain A? levels. Early in disease pathogenesis, soluble A? monomer aggregates into soluble A? oligomers and insoluble A? plaques, both of which cause synaptic dysfunction and kill neurons under certain conditions. In 2009, we were the first to demonstrate a diurnal fluctuation in brain interstitial fluid (ISF) A? levels in APP transgenic mice, with highest levels during wakefulness and lowest during sleep. In humans, CSF A? levels have a similar diurnal fluctuation as mice and recent studies demonstrated that a single night of sleep deprivation acutely increases CSF A?. Numerous groups have independently replicated that sleep, A?, and AD are intimately linked; however, the cellular mechanisms underlying this link remain unclear. Synaptic activity appears to be one mediator driving the diurnal fluctuation in A? during the sleep-wake cycle. Our preliminary data demonstrate that the neuropeptide orexin/hypocretin drives the sleep-wake fluctuation in A?. Orexin is produced exclusively within the hypothalamus and is released throughout the brain to induce wakefulness. Our data suggest that orexin initiates a signaling cascade that ultimately increases brain A? levels. We propose that this signaling pathway is the cellular link between the sleep-wake cycle and A? regulation. Interestingly, orexin is co-released with another neuropeptide, dynorphin. Our data demonstrate that dynorphin has the opposite effect on A? levels from orexin; dynorphin A and a selective agonist for the dynorphin kappa-opioid receptor (KOR) subtype suppresses brain ISF A? levels, whereas a KOR antagonist increases A? in vivo. We hypothesize that orexin and dynorphin neuropeptides act in concert to regulate A? levels in opposite directions. Our goal is to define the signaling pathways that underlie this regulation, as well as determine how the receptors interact to influence A? levels and pathology. We will use in vivo microdialysis to define this interaction in living mice within a normal sleep-wake pattern so our findings are physiologically in context. We will also determine how chronic modulation of orexin and dynorphin signaling impacts A? pathology and behavior deficits. These studies will provide insight into how sleep relates to disease risk and suggest new therapeutic avenues. Orexin and dynorphin are targets of several FDA-approved drugs. Admittedly, orexin receptors and dynorphin opioid receptors have complicated biological and behavioral effects. Our hope is that understanding the signaling biology and interaction in depth will provide a means to use approved agents, or novel ones, in the setting of AD.